Investigation of BBE-like enzymes in Neurospora crassa

Description

Fungi are recognized for their ability to generate a wide variety of natural compounds serving various purposes. These natural products are usually synthesized from simpler activated building blocks that can undergo a series of enzymatic modifications, leading to structurally diverse and complex molecules. Redox reactions play a pivotal role in determining the biological activity of natural product.1 In order to catalyze the desired redox reactions, oxidoreductases, like flavin-dependent enzymes, are required.2 To study the involvement of flavoenzymes in the biosynthesis of natural products, the filamentous fungus N. crassa, the red bread mold, was chosen as a model organism. The flavoproteom of N. crassa is encoded by 201 genes corresponding to 2% of the genome.1 Nearly two-thirds of the genes encode flavoenzymes that play a role in primary metabolism. The remaining genes (ca. 68) encode flavoenzymes of yet unknown function. Among these genes, seventeen encode BBE-like enzymes. Phylogenetic analysis indicated that these BBE-like enzymes can be classified into four clades: alcohol oxidases, EncM-like enzymes, carbohydrate oxidases, and cytokinin oxidases. Seven of these BBE-like enzymes were chosen for heterologous production and characterization. Here, we report the successful production and crystallization of NcBBE-like 14. The enzyme is a thermostable oxidase with a redox potential of +57 mV and an oxidative rate of 4.0 x 105 M-1 s-1. Like other BBE-like enzymes, it shares the vanillyl oxidase fold and comprises three domains: a substrate- and FAD-binding domain and a BBE domain (Figure 1). The crystal structure also reveals a monocovalent linkage between the 8-α position of the cofactor and the enzyme’s histidine residue H100.

Period	15 Jul 2024 → 24 Jul 2024
Event title	21st International Flavins and Flavoproteins Symposium
Event type	Conference
Location	Atlanta, United StatesShow on map
Degree of Recognition	International